Looking for breakthrough ideas for innovation challenges? Try Patsnap Eureka!

A fully human neutralizing antibody against h7n9 virus

A technology of antibodies and viruses, applied in the fields of cellular immunology and molecular biology, can solve problems such as uncontrollable results, unoptimistic development prospects, and limited clinical applications

Active Publication Date: 2020-05-05
JIANGSU PROVINCIAL CENT FOR DISEASE PREVENTION & CONTROL
View PDF3 Cites 0 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

At present, the research on monoclonal neutralizing antibodies against H7N9 influenza at home and abroad is mostly mouse-derived monoclonal antibodies. The clinical application of this mouse-derived monoclonal antibody is limited due to the immune reaction of mouse proteins.
Although the humanized antibody that uses genetic engineering technology to humanize the mouse-derived monoclonal antibody is currently the dominant monoclonal antibody product market, it still retains the mouse-derived variable region sequence, coupled with human The source process is complicated and the result is uncontrollable, and its development prospects are not optimistic

Method used

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
View more

Image

Smart Image Click on the blue labels to locate them in the text.
Viewing Examples
Smart Image
  • A fully human neutralizing antibody against h7n9 virus
  • A fully human neutralizing antibody against h7n9 virus
  • A fully human neutralizing antibody against h7n9 virus

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0108] Example 1 Preparation of fully human monoclonal neutralizing antibody mAb635 against H7N9 virus

[0109] 1. Memory B cell sorting

[0110] 1.1 Isolation of PBMCs

[0111] Three samples of peripheral anticoagulant blood from H7N9 convalescent patients were collected, and PBMCs from peripheral blood were separated by Ficoll density gradient centrifugation and stored in liquid nitrogen.

[0112] 1.2 Acquisition of memory B cells

[0113] The purified HA protein was labeled with PE and APC fluorescent dyes, and then mixed with Anti-CD3-Cy7APC, Anti-CD14-FITC, Anti-CD20-Cy7PE, Anti-CD19-PE-CF594, Anti-IgG-BV421, Anti-IgM -Cy55 fluorescently labeled antibodies were mixed in corresponding proportions. Take out PBMC cells from liquid nitrogen, wash with PBS after thawing, first stain the dead cells with AQUA staining solution, and then incubate with the above-mentioned mixed antibody on ice to bind. Unbound antibodies were washed by centrifugation at 800g, washed with PBS a...

Embodiment 2

[0130] Example 2 Detection of mAb635 Antibody Binding Specificity

[0131] Detection of binding of purified antibody to H7N9 virus HA protein by immunoblotting

[0132] 1. Steps:

[0133] (1) The HA protein of the purified H7N9 virus was subjected to SDS-PAGE, and the HA protein of the purified H3N2 virus was used as a control at the same time. After the electrophoresis was completed, the protein in the gel was transferred to a PVDF membrane, and the PVDF membrane was 5% skimmed milk 4 Block overnight at ℃, then add 50μg of purified antibody, incubate at 37℃ for 2h, and then elute with PBST;

[0134] (2) Add anti-human IgG secondary antibody at 1:2000, incubate at 37°C for 1 hour, and then elute with PBST;

[0135] (3) PVDF membrane plus chemiluminescent liquid is exposed in a dark room.

[0136] 2. Results

[0137] The result is as image 3 As shown, the purified mAb635 antibody specifically binds to the H7N9 virus HA protein, image 3 "1" represents the lane where the ...

Embodiment 3

[0138] Example 3 Hemagglutination inhibition experiment of mAb635 antibody

[0139] 1. Treatment of serum and antibody

[0140] A positive control serum is the serum of a patient in the recovery period of H7N9 virus infection, and a negative control serum is the serum of a healthy adult. Add 3 times the volume of RDE to the serum and antibody, incubate at 37°C for 16-18 hours, inactivate at 56°C for 30 minutes before use, add 6 times the volume of PBS to the serum, and the final concentration is 1:10, without adding the antibody, the final concentration is 1:1: 4.

[0141] 2. Preparation of 1% turkey red blood cells

[0142] Inhale 2ml of Alfred's solution in the syringe in advance, and take 2ml of venous blood from the root of the turkey wing to form a 4ml whole blood mixture. Add 4ml of blood to a 50ml centrifuge tube, add cold PBS solution to the centrifuge tube to 50ml, mix gently and centrifuge at 2000rpm for 5 minutes to remove the supernatant suspension, then wash pa...

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to View More

PUM

No PUM Login to View More

Abstract

The invention discloses a fully-human-derived H7N9 virus resistant monoclonal neutralizing antibody or an antigen combining segment thereof. The neutralizing antibody can specifically combine with H7N9 virus and can effectively neutralize virus activity. The antibody can be used for preventing, treating and detecting the H7N9 virus and has wide application prospect clinically.

Description

technical field [0001] The invention belongs to the fields of cellular immunology and molecular biology, and relates to a fully human monoclonal antibody, in particular to a fully human monoclonal neutralizing antibody against H7N9 virus. In addition, the present invention also relates to the preparation method and application of the neutralizing antibody. Background technique [0002] The H7N9 virus belongs to the Orthomyxoviridae family. It is a segmented RNA virus with a single negative strand and an envelope. The genome is divided into 8 segments. The virus is spherical, about 100nm in size, and consists of a capsid and an envelope. Eight viral RNA segments are covered by NP to form a helically symmetrical viral capsid. The envelope comes from the host cell membrane, and HA and NA protein spikes are embedded on the surface. HA is chimerized on the surface of the envelope in the form of a trimer. It is the main antigenic component of influenza virus, can agglutinate red...

Claims

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to View More

Application Information

Patent Timeline
no application Login to View More
Patent Type & Authority Patents(China)
IPC IPC(8): C07K16/10C12N15/13A61K39/42A61P31/14G01N33/569
CPCA61K39/00C07K16/1018C07K2317/24C07K2317/565C07K2317/76
Inventor 朱凤才张黎李靖欣孟繁岳胡月梅
Owner JIANGSU PROVINCIAL CENT FOR DISEASE PREVENTION & CONTROL
Who we serve
  • R&D Engineer
  • R&D Manager
  • IP Professional
Why Patsnap Eureka
  • Industry Leading Data Capabilities
  • Powerful AI technology
  • Patent DNA Extraction
Social media
Patsnap Eureka Blog
Learn More
PatSnap group products

Browse by: Latest US Patents, China's latest patents, Technical Efficacy Thesaurus, Application Domain, Technology Topic, Popular Technical Reports.

© 2024 PatSnap. All rights reserved.Legal|Privacy policy|Modern Slavery Act Transparency Statement|Sitemap|About US| Contact US: help@patsnap.com